Background: Diagnostic error is a global patient safety priority. Objectives: To estimate the incidence, origins and avoidable harm of diagnostic errors in English general practice. Diagnostic errors were defined as missed opportunities to make a correct or timely diagnosis based on the evidence available (missed diagnostic opportunities, MDOs). Method: Retrospective medical record reviews identified MDOs in 21 general practices. In each practice, two trained general practitioner reviewers independently conducted case note reviews on 100 randomly selected adult consultations performed during 2013-2014. Consultations where either reviewer identified an MDO were jointly reviewed. Results: Across 2057 unique consultations, reviewers agreed that an MDO was possible, likely or certain in 89 cases or 4.3% (95% CI 3.6% to 5.2%) of reviewed consultations. Inter-reviewer agreement was higher than most comparable studies (Fleiss' kappa=0.63). Sixty-four MDOs (72%) had two or more contributing process breakdowns. Breakdowns involved problems in the patient-practitioner encounter such as history taking, examination or ordering tests (main or secondary factor in 61 (68%) cases), performance and interpretation of diagnostic tests (31; 35%) and follow-up and tracking of diagnostic information (43; 48%). 37% of MDOs were rated as resulting in moderate to severe avoidable patient harm. Conclusions: Although MDOs occurred in fewer than 5% of the investigated consultations, the high numbers of primary care contacts nationally suggest that several million patients are potentially at risk of avoidable harm from MDOs each year. Causes of MDOs were frequently multifactorial, suggesting the need for development and evaluation of multipronged interventions, along with policy changes to support them.